L0301P17 - RNA Synthesis
__TOC__ RNA Types *messenger RNA - mRNA **code for proteins **can be of many different sizes depending on the protein being produced *ribosomal RNA - rRNA **contributes to the structure and function of ribosomes *transfer RNA - tRNA **carry amino acids and act as adaptor molecules allowing decoding of mRNA to protein Structure and Function *hydroxyl group on ribose and single-stranded-ness makes it unstable within the cell = short-lived/transient *complementarity - bases can still form pairs = folding to produce tRNA structure *complexity = perfect for formation of higher order structures RNA Polymerases *prokaryotes: **one type of RNA polymerase that transcribes mRNA, tRNA and rRNA *eukaryotes: **three types: I, II, III **RNA polymerase II makes mRNA **RNA polymerases can make errors for every 104 to 105 bases as it is not proofread **errors do not cause many problems as mRNA is relatively short-lived Transcription *RNA transcribed from DNA *one of the two strands for a gene will be used to make RNA = template strand *the other strand that is not used to make RNA is called the non-coding strand *newly synthesised RNA would have the same bases as the non-coding strand except uracil replaces thymine *different genes in the same DNA molecule may not use the same strand as the template *RNA polymerase uses DNA as a template to make RNA Initiation *DNA polymerase recognises and binds tightly to the promoter sequence on DNA **promoter sequence - a region of DNA that occurs upstream of the coding region *this binding can influenced by other proteins and initiates RNA synthesis *promoter sequence directs the RNA polymerase as to: **which of the two strands is the template **in what direction it should move Elongation *RNA polymerase unwinds the DNA about 10 bp at a time exposing the template *RNA polymerase reads the template beginning at the initiation site in the 3’ to 5’ direction *RNA is synthesised in the 5’ to 3’ direction so that the transcript is antiparallel to the DNA template strand *exiting RNA transcript falls away from the DNA allowing it to be rewound into the double helix *energy for synthesis comes from removal and breakdown of the pyrophosphate (two phosphates) group from each nucleoside triphosphate (NTP) added Termination *particular base sequences in the DNA specify termination *in eukaryotes **fairly complex protein is required **does not occurs until a sequence signalling polyadenylation (series of A’s added) has been transcribed RNA Processing Chemical Modifications *after transcription, pre-mRNA is altered by: **addition of a methyl-G cap at the 5’ end **addition of a poly-A tail at the 3’ end **after cleavage at AAUAAA sequence *tail may vary in length (affects stability) *modifications are important for stability of the RNA and competence to be translated Splicing *introns are removed from mRNA precursor by spliceosomes *small nuclear ribonucleoprotein particles (snRNPs) bind to pre-mRNA as soon as it is transcribed *this allows linkage of all the exons into one continuous sequence of mature mRNA *extremely precise and accurate process otherwise will affect the proteins produced Genetic Code *refers to triplets of bases (codons) as which mRNA is read *64 codons to code for 20 amino acids and start and stop signals: **AUG: methionine = start of translation **UAA, UAG, UGA: no a.acid = terminate *is redundant - many amino acids have more than one codon *is not ambiguous - each codon is always assigned to only one amino acid *is nearly universal - applies to all species, minor variation in mitochondria, chloroplasts Expression of the Eukaryotic Genome *eukaryotic DNA is separated form the cytoplasm by being contained in the nucleus *initial mRNA transcript must be bodied before export to the cytoplasm Control of Gene Expression *in eukaryotes, can be controlled at the transcriptional, post-transcriptional, translational and post-translational levels Remodelling of Chromatin *first level of regulation - to allow access of RNA synthesis enzymes to the DNA *the negatively charged DNA must be first partially detached from the positively charged histone and unwound *attraction between histone and DNA can be weakened by acetylation Selective Transcription *results from transcription factors (proteins) binding to regulatory regions on DNA **some bind to promoter region before RNA polymerase can bind **some work with RNA polymerase II to transcribe mRNA *activators **CRP-cAMP complex binds to the promoter enhancing efficiency of the binding of RNA polymerase **can be metabolite-dependent *repressors **some bind to operators blocking RNA polymerase from binding to the promoter (lac operon) **others bind until inducer binds to it and inactivated it (allolactose) **simultaneous control of widely separated genes is possible through TFs that bind to common sequences in their promoters Post Transcriptional Control *~50% of eukaryotic genes have several exons and alternative splicing can occur to produce different (possibly related) proteins *stability of mRNA in the cytoplasm can be regulated by the binding of proteins. *specific AU-rich sequences mark some mRNAs for rapid breakdown by a ribonuclease complex (exosome) RNA Involvement - microRNAs *short, 19-25 nucleotide, non-coding RNA *when combined with particular proteins, they bind to complementary sequences found in some mRNAs and prevents their translation and facilities degradation *de-regulated in some cancer cells (over-expressed) *exact number of miRNA genes is not known **may regulate 10-30% of human genes Translational Control *often little correlation between the amount of mRNA and the amount of protein produced, thus factors must be affecting these amounts after mRNA is made *binding of repressor proteins to the mRNA to inhibit translation **bind to noncoding regions of mRNA (known as the riboswitch) and block translation by preventing it from binding to a ribosome Post-Translational Control *i.e. regulation of protein longevity *most gene products (proteins) are modified after translation *regulating the lifetime of a protein is a way to control its actions. *one common method known: ubiquitination *proteins identified for breakdown are often linked to a 76-amino acid protein, ubiquitin *other proteins detect proteins that are no longer required and attach ubiquitin chains onto them, which acts as a label *protein–ubiquitin complex then binds a complex called a proteasome *inside the proteasome, ATP used to cut off the ubiquitin (for recycling) and unfold its targeted protein *proteases digest the protein into small peptides and amino acids